Tumors are potentially immunogenic; however, they fail to spontaneously induce an immune response capable of rejecting the tumor. A major reason is that the tumor microenvironment lacks adequate innate immune activation required to initiate strong adaptive anti-tumor immunity. Plasmacytoid dendritic cells (pDC) comprise a dendritic cell subset highly specialized in sensing microbial nucleic acids via intracellular Toll-like receptors. During viral infection, pDC accumulate in infected tissues and are activated by viral nucleic acids to produce large amounts of type I IFNs and generate protective immunity against the virus through activation of myeloid DCs, T cells, and NK cells. Tumors also contain pDCs, but do not provide the molecular signals to activate pDCs. Although tumors contain high concentrations of self-DMA released in the extracellular environment as a result of the increased turnover and tumor cell death, it is clear that pDCs, while activated by viral nucleic acids, are normally not able to sense tumor-derived DMA and are thereby unable to initiate a strong innate immune response. We recently found that pDC can, in fact, sense and respond to self-DMA when combined with an endogenous peptide called LL37. LL-37 can bind self-DMA fragments released by dying cells to form aggregates and condensed structures that are delivered to and retained within early endosomes of pDCs. In these intracellular compartments, LL37/DNA can interact with TLR9 to trigger robust type I IFN production similarly to viral DMA. Because tumors release large amounts of self-DMA and contain pDCs but do not express LL37, our hypothesis is that exogenous LL37 can be used to target tumor-derived self-DMA and convert it into a danger signal that triggers pDC activation and type I IFN production at the tumor site. This will then induce T-cell mediated immunity against the tumor by the same mechanism by which anti-viral immune responses are induced. Specific Aim 1 will determine whether LL37 can convert self-DMA released by dying tumor cells from being immunologically inert into a trigger of pDC activation to produce type I IFNs. Specific Aim 2 will evaluate whether LL37 mixed with dying tumor cells ex-vivo and injected in-vivo as a vaccine can induce effective T cell-mediated anti-tumor immunity. Specific Aim 3 will assess whether dying tumor cells releasing self-DMA can be targeted by intratumoral injection or systemic administration of LL37 to induce innate immune activation at the tumor site leading to effective anti-tumor immunity. These studies may lead to the design of future clinical trials utilizing LL37 in cancer patients with diffusely metastatic disease.